An effective vaccine for the prevention of HIV-1 infection will likely be one that induces a balanced immune response, stimulating both neutralizing antibody and cellular immune responses. The principal HIV- 1 antigen to which virtually all neutralizing antibody is directed is the envelope glycoprotein (Env) of the virus. The biologically relevant form of ENV is oligomeric, and the location of the Env at the surface of the virus render it an important target for vaccine development. To date, most vaccine-induced neutralizing antibody responses to first-generation monomeric Env antigens have been limited in both potency and cross- reactivity among primary isolates of HIV-1, and this is likely do to the type of Env used in the vaccine. Thus, it is important to understand the determinants in Env which stimulate the production of antibodies that are broadly reactive. We have found that immunization with soluble, oligomeric Env effectively generates a more broadly cross-reactive antibody response, and our studies have indicated the importance of further investigating oligomeric Env as an immunogen. One way neutralizing antibody antibodies could work to perturb some metastable property of Env required for fusion. It is likely that these antibodies will react with highly conserved epitopes dependent on oligomeric structure and critical for its function, structures from which the myriad of HIV-1 isolates cannot evolve to escape. Such antibodies would possess broadly cross-reactive potential. Finally, since we know it is possible to elicit neutralizing antibody, then new methods to deliver these antigens that will bolster the level and longevity these responses will be important. Combining these goals with the Venezuelan equine encephalitis virus (VEE) vaccine studiers, a system known to stimulate CTL activity, will yield further information for an overall program aimed at the development of an efficacious vaccine for the prevention of HIV-1 infection. The overall goal of this project is to fully evaluate oligomeric Env as an immunogen in eliciting a neutralizing antibody response. Specifically, we will: 1) Develop and characterize a variety of soluble oligomeric Env immunogens from representative clade B', C and E primary HIV-1 isolates, and from the relevant isolate R2; 2) Use oligomeric gp140s of R2 and candidate B', C and E isolates as immunogens with and without a novel encapsulated delivery method in mice, rabbits and macaques; and 3) Determine the protective efficacy of clade-specific immunity induced by purified soluble oligomeric gp140 as a protein-based subunit immunogen alone and in VEE replicon immunized macaques.